PROJECT SUMMARY Preclinical and clinical development of molecular targeted drugs with radiation therapy (RT) and chemotherapy are critically important for improving the outcomes of patients with hard-to-treat cancers. However, a huge body of preclinical drug/RT studies has not translated into an adequate number of successful radiation oncology trials. Major contributing factors include poor reproducibility of preclinical data, insufficient preclinical modeling of inter-tumoral genomic heterogeneity that influences treatment sensitivity in the clinic, and reliance on tumor growth delay instead of local tumor control (TCD50) endpoints. There exists an urgent need to overcome these barriers to successful clinical translation of targeted chemoradiosensitizers. We propose to establish an integrated in-vitro/in-vivo pipeline for chemoradiosensitizing targeted drugs that are biomarker-correlated and appropriately validated, so that subsequent clinical drug/RT trials in patients with hard-to-treat cancers will have a substantially higher probability of success than in the past. To achieve this, we propose 3 Specific Aims. First, by leveraging the unique expertise and resources that the Genomics of Drug Sensitivity in Cancer project (Massachusetts General Hospital & UK Wellcome Trust Sanger Institute) and the German Cancer Research Center/Cancer Consortium (DKFZ/DKTK) offer, we will conduct a robotic high-throughput screen of cancer cell lines grown in an extracellular-matrix (ECM) based 3D format to better mimic in-vivo growth conditions. We propose to screen about half of the current CTEP portfolio (30 drugs) combined with fractionated irradiation across an initial panel of 100 annotated cell lines selected to represent clinically relevant inter-tumoral genomic variation. Second, we propose a systematic and stepwise validation/refinement process to nominate CTEP drugs that have the highest likelihood to succeed in animal testing. This will include 3D colony formation assays, addition of disease-specific chemotherapy, confirmation of pharmacodynamic responses, target/biomarker validation, and integration of patient-derived cell lines and xenografts. Third, we will evaluate the chemoradiosensitizing effects of the most promising CTEP drugs in-vivo by relying on TCD50 assays in mouse xenograft models with/without biomarker and utilizing clinically relevant RT fractionation (30 fractions/6 weeks). These experiments again will leverage special DKFZ/DKTK capability. The proposed studies are directly relevant to the objectives of the underlying FOA, as we will, in close collaboration with investigators within and outside the consortium, accelerate the pace at which targeted chemoradiation treatments with greater efficacy are identified. This undertaking will be greatly facilitated by the integration of a foreign project component where DKFZ/DKTK investigators contribute special expertise in 3D ECM and TCD50 assays at low budget cost, which will directly and disproportionally benefit the NCI and the radiation oncology community in the United States. By integrating extensive expertise in pharmacogenomics and radiation biology, the inter-disciplinary investigator team is uniquely poised to help transform the preclinical discovery process for chemoradiosensitizing targeted drugs with accompanying biomarkers.